The device and method in accordance with the constant pressure regulation of micro droplet PCR in microfluidic chips are developed to explore the optimization way for the micro droplet movement, fragmentation, and bubble generation in microfluidic chips in existing digital PCR technology. In the developed device, an air source device is adopted to regulate the pressure in the chip, such that micro droplet generation and PCR amplification without bubbles can be achieved. In 3 min, the sample in 20ul will be distributed into nearly 50,000 water-in-oil droplets exhibiting a diameter of about 87μm, and the micro droplet will be subjected to a close arrangement in the chip without air bubbles. The device and chip are adopted to quantitatively detect the human gene . As indicated by the experimental results, a good linear relationship exists between detection signal and DNA concentration ranging from 101~ 105 copies /μL (R2=0.999). The micro droplet PCR devices based on constant pressure regulation chips exhibit a wide variety of advantages (e.g., achieving high pollution resistance, micro droplet fragmentation and integration avoidance, reducing human interference, and standardizing results). Thus, micro droplet PCR devices based on constant pressure regulation chips have promising applications in nucleic acid quantification.